Cloning of three type III polyketide synthases and formation of polyketides in recombinant using cinnamic acid analogs as substrates.

is an actinorhizal plant that coexists with the nitrogen-fixing actinomycete via nodules. It produces a variety of polyketides, including flavonoids, stilbenoids, and diarylheptanoids. These compounds have beneficial biological activities. Plant polyketides are produced by type III polyketide synthases (PKSIII). In this study, three PKSIIIs (AsPKSIII1, AsPKSIII2, and AsPKSIII3) predicted from next-generation sequencing analysis of seedling RNA were amplified and cloned. Phylogenetic tree analysis classified AsPKSIII2 and AsPKSIII3 into the chalcone synthase (CHS) group, whereas AsPKSIII1 was not classified into this group. We attempted to produce polyketides by adding cinnamic acid analogs to the culture medium of in which the respective PKSIII gene and the acetyl-CoA carboxylase (ACC) and 4-coumarate: CoA ligase (4CL) genes were simultaneously recombined. AsPKSIII1 is an enzyme that condensed only one molecule of malonyl-CoA to cinnamoyl-CoAs. In contrast, AsPKSIII2 and AsPKSIII3 produced chalcones as shown in a phylogenetic tree analysis, but also produced triketide pyrone. The ratio of these products differed between the two enzymes. We determined the gene and amino acid sequences as well as the substrate specificities of the two enzymes involved in flavonoid production and one enzyme potentially involved in diarylheptanoid production in .